The object of the proposed research is to gain an understanding of the biochemical mechanism responsible for the synthesis of mRNA in mammalian cells. We will make use of an in vitro transcription system to study the requirements of the various steps involved in the synthesis of a mature mRNA molecule. A number of deletion and point mutations located around the apparent start-site of a well characterized viral transcript (the human adenovirus major late transcript) will be analyzed in detail. The affects of these mutations on transcription initiation will be determined. The relationship between RNA chain initiation and capping will be studied. We will optimize and modify the in vitro system so that the requirements for RNA splicing, 3' end formation and polyadenylation can be elucidated. This work will involve the synthesis of new DNA templates, by in vitro recombinant DNA technologies, to detect these activities.